1. Field of the Invention
The invention relates to compositions comprising a retinoid X receptor agonist and an agent capable of activating protein kinase A. The invention also relates to methods of treating hyperproliferative diseases by administering a retinoid X receptor agonist and an agent capable of activating protein kinase A.
2. Related Art
Retinoids and Receptors
A number of studies have demonstrated that retinoids (vitamin A derivatives) are essential for normal growth, vision, tissue homeostasis, reproduction and overall survival (for reviews and references, see Sporn et al., The Retinoids, Vols. 1 and 2, Sporn et al., eds., Academic Press, Orlando, Fla. (1984)).
Except for those involved in visual perception (Wald, G. et al., Science 162:230-239 (1968)), the molecular mechanisms underlying the highly diverse effects of retinoids have until recently remained obscure. The discovery of nuclear receptors for retinoic acid (RA) (Petkovich et al., Nature 330:444-450 (1987); Giguxc3xa8re et al., Nature 330:624-629 (1987)) has greatly advanced the understanding of how the retinoids may exert their pleiotropic effects (Leid, M., et al., TIBS 17:427-433 (1992); Linney, E., Current Topics in Dev. Biol. 27:309-350 (1992)). It is thought that the effects of the RA signal are mediated through two families of receptorsxe2x80x94the RAR family and RXR familyxe2x80x94which belong to the superfamily of ligand-inducible transcriptional regulatory factors that include steroid/thyroid hormone and vitamin D3 receptors (for reviews, see Leid, M., et al., TIBS 17:427-433 (1992); Chambon, P., Semin. Cell Biol. 5:115-125 (1994); Chambon, P., FASEB J. 10:940-954 (1996); Giguere, V., Endocrinol. Rev. 15:61-79 (1994); Mangelsdorf, D. J., and Evans, R. M., Cell 83:841-850 (1995); Gronemeyer, H., and Laudet, V., Protein Profile 2:1173-1236 (1995)).
Receptors belonging to the retinoic acid receptor family (RARxcex1, xcex2 and xcex3 and their isoforms) are activated by both all-trans- and 9-cis-RA (Leid, M., et al., TIBS 17:427-433 (1992); Chambon, P., Semin. Cell Biol. 5:115-125 (1994); Dolle, P., et al., Mech. Dev. 45:91-104 (1994)). Unlike the RARs, members of the retinoid X receptor family (RXRxcex1, xcex2 and xcex3) are activated exclusively by 9-cis-RA (Chambon, P., Semin. Cell Biol. 5:115-125 (1994); Dollxc3xa9, P., et al., Mech. Dev. 45:91-104 (1994); Linney, E., Current Topics in Dev. Biol. 27:309-350 (1992); Leid, M., et al., TIBS 17:427-433 (1992); Kastner et al., In: Vitamin A in Health and Disease, R. Blomhoff, ed., Marcel Dekker, New York (1993)).
Nuclear receptors (NRs) are members of a superfamily of ligand-inducible regulatory factors that include receptors for steroid hormones, thyroid hormones, vitamin D3 and retinoids (Leid, M., et al., Trends Biochem. Sci. 17:427-433 (1992); Leid, M., et al., Cell 68:377-395 (1992); and Linney, E. Curr. Top. Dev. Biol., 27:309-350 (1992)). NRs exhibit a modular structure which reflects the existence of several autonomous functional domains. Based on amino acid sequence similarity between the chicken estrogen receptor, the human estrogen and glucocorticoid receptors, and the v-erb-A oncogene, Krust, A., et al. (EMBO J. 5:891-897 (1986)) defined six regionsxe2x80x94A, B, C, D, E and Fxe2x80x94which display different degrees of evolutionary conservation among various members of the nuclear receptor superfamily. The highly conserved region C contains two zinc fingers and corresponds to the core of the DNA-binding domain (DBD), which is responsible for specific recognition of the cognate response elements. Region E is functionally complex, since in addition to the ligand-binding domain (LBD), it contains a ligand-dependent activation function (AF-2) and a dimerization interface. An autonomous transcriptional activation function (AF-1) is present in the non-conserved N-terminal A/B regions of the steroid receptors. Interestingly, both AF-1 and AF-2 of steroid receptors exhibit differential transcriptional activation properties which appear to be both cell type and promoter context specific (Gronemeyer, H., Annu. Rev. Genet. 25:89-123 (1991)).
It has been shown that activation of RA-responsive promoters likely occurs through RAR/RXR heterodimers rather than through homodimers (Yu, V.C., et al., Cell 67:1251-1266 (1991); Leid, M., et al., Cell 68:377-395 (1992b); Durand et al., Cell 71:73-85 (1992); Nagpal, S., et al., Cell 70:1007-1019 (1992); Zhang, X. K., et al., Nature 355, 441-446 (1992); Kliewer et al., Nature 355:446-449 (1992); Bugge et al., EMBO J. 11:1409-1418 (1992); Marks et al., EMBO J. 11:1419-1435 (1992); Yu, V. C. et al., Cur. Op. Biotech. 3:597-602 (1992); Leid, M., et al., TIBS 17:427-433 (1992); Laudet and Stehelin, Curr. Biol. 2:293-295 (1992); Green, S., Nature 361:590-591 (1993)). The RXR portion of these heterodimers has been proposed to be silent in retinoid-induced signaling (Kurokawa, R., et al., Nature 371:528-531 (1994); Forman, B. M., et al., Cell 81:541-550 (1995); Mangelsdorf, D. J., and Evans, R. M., Cell 83:835-850 (1995); Vivat, V. et al., EMBO J. 16:5697-5709 (1997)) but conflicting results have been reported as far as the ligand-binding ability of RXR in heterodimers is concerned (Kurokawa, R., et al., Nature 371:528-531 (1994); Chen, J.-Y. et al., Nature 382:819-822 (1996); Kersten, S. et al., Biochem. 35:3 816-3824 (1996); Chen, Z. et al., J. Mol. Bio. 275:55-65 (1998); Li, C. et al., Proc. Natl. Acad. Sci. USA 94:2278-2283 (1997). The results of these and of genetic studies strongly suggest that RAR/RXR heterodimers are indeed functional units that transduce the RA signal in vivo (Chambon, P., Semin. Cell Biol. 5:115-125 (1994); Kastner, P. et al., Cell 83:859-869 (1995); Mascrez, B. el al., Development 125:4691-4707 (1998)). Thus, the basis for the highly pleiotropic effect of retinoids may reside, at least in part, in the control of different subsets of retinoid-responsive promoters by cell-specifically expressed heterodimeric combinations of RAR/RXR subtypes (and isoforms), whose activity may be in turn regulated by cell-specific levels of all-trans- and 9-cis-RA (Leid, M., et al., TIBS 17:427-433 (1992)).
The RXR receptors may also be involved in RA-independent signaling. For example, the observation of aberrant lipid metabolism in the Sertoli cells of RXRxcex2+ mutant animals suggests that functional interactions may also occur between RXRxcex2 and the peroxisomal proliferator-activated receptor signaling pathway (WO 94/26100; Kastner, P., et al., Genes and Dev. 10:80-92 (1996)).
Therapeutic Uses of Retinoids
Overview
As retinoic acid is known to regulate the proliferative and differentiative capacities of several mammalian cell types (Gudas, L. J., et al., In: THE RETINOIDS, 2nd ed., Sporn, M. B., et al., eds., New York: Raven Press, pp. 443-520 (1994)), retinoids are used in a variety of chemopreventive and chemotherapeutic settings. The prevention of oral, skin, head and neck cancers in patients at risk for these tumors has been reported (Hong, W. K., et al., N. Engl. J. Med. 315:1501-1505 (1986); Hong, W. K., et al., N. Engl. J. Med. 323:795-801 (1990); Kraemer, K. H., et al., N. Engl. J. Med. 318:1633-1637 (1988); Bollag, W., et al., Ann. Oncol. 3:513-526 (1992); Chiesa, F., et al., Eur. J. Cancer B. Oral Oncol. 28:97-102 (1992); Costa, A., et al., Cancer Res. 54:Supp. 7,2032-2037(1994)). Retinoids have also been used to treat squamous cell carcinoma of the cervix and the skin (Verma, A. K., Cancer Res. 47:5097-5101 (1987); Lippman S. M., et al., J. Natl Cancer Inst. 84:235-241 (1992); Lippman S. M., et al., J. Natl Cancer Inst. 84:241-245 (1992)) and Kaposi""s sarcoma (Bonhomme, L., et al., Ann. Oncol. 2:234-235 (1991)), and have found significant use in the therapy of acute promyelocytic leukemia (Huang, M. E., et al., Blood 72:567-572 (1988); Castaigne, S., et al., Blood 76:1704-1709 (1990); Chomienne, C., et al., Blood 76:1710-1717 (1990); Chomienne, C., et al., J. Clin. Invest. 88:2150-2154 (1991); Chen Z., et al., Leukemia 5:288-292 (1991); Lo Coco, F., el al., Blood 77:1657-1659 (1991); Warrell, R. P., et al., N. Engl. J. Med. 324:1385-1393 (1991); Chomienne, C., et al., FASEB J. 10:1025-1030 (1996)). Retinoids are also used to treat hyperproliferative skin disorders such as psoriasis. 13-cis retinoic acid (isotretinoin) is commonly used as a dermatologic drug.
Acute Promyelocytic Leukemia (APL)
A balanced chromosomal translocation, t(15; 17), has been identified in most acute promyelocytic leukemia (APL) cells (Larson, A. R., et al., Am. J. Med. 76:827-841 (1984)). The breakpoint for this translocation occurs within the second intron of the RARxcex1 gene (Alcalay, M. D., et al., Proc. Natl. Acad. Sci. USA 88:1977-1981 (1991); Chang, K. S., el al., Leukemia 5:200-204 (1991); Chen, S., et al., Blood 78:2696-2701 (1991) and within two loci of the gene encoding the putative zinc finger transcription factor PML (Goddard, A., et al., Science 254:1371-1374(1991)). This reciprocal t(15;17) translocation leads to the generation of a PML-RARxcex1 fusion protein which is co-expressed with PML and RARxcex1 in APL cells (see Warrell, R. P., et al., N. Engl. J. Med. 329:177-189 (1993); Grignxc3xa1ni, F., et al., Blood 83:10-25 (1994); Lavau, C., and Dejean, A., Leukemia 8:1615-1621 (1994); de Thxc3xa9, H., FASEB J. 10:955-960 (1996)). The PML-RARxcex1 fusion is apparently responsible for the differentiation block at the promyelocytic stage, since (i) it is observed in nearly all APL patients (Warrell, R. P., et al., N. Engl. .J. Med. 329:177-189 (1993); Grignxc3xa1ni, F., el al., Blood 83:10-25 (1994); Lavau, C., and Dejean, A., Leukemia 8:1615-1621 (1994)), (ii) it inhibits myeloid differentiation when overexpressed in U937 or HL60 myeloblastic leukemia cells (Grignani, F., el al., Cell 74:423-431 (1993)), and (iii) complete clinical remission due to differentiation of the leukemic cells to mature granulocytes upon treatment with all-trans retinoic acid (ATRA) is tightly linked to PML-RARxcex1 expression (Warrell, R. P., et al., N. Engl. J. Med. 324:1385-1393 (1991); Lo Coco, R., et al., Blood 77:1657-1659 (1991); Chomienne, C., et al., FASEB J. 10:1025-1030 (1996)). Multiple studies have addressed the possible impact of PML-RARxcex1 fusion protein formation on cell proliferation (Mu, X.M., et al., Mol. Cell. Biol. 14:6858-6867 (1994)) and apoptosis (Grignani, F., et al., Cell 74:423-431 (1993)), AP1 transrepression (Doucas, V., et al., Proc. Natl. Acad. Sci. USA 90:9345-9349(1993)), and vitamin D3 signaling (Perez, A., et al., EMBO J. 12:3171-3182 (1993)), but the mechanism(s) by which PML-RARxcex1 blocks myeloid cell maturation has remained elusive. Consistent with the aberrant nuclear compartmentalization of PML-RARxcex1, which adopts the xe2x80x9cPML-typexe2x80x9d location upon RA treatment (Dyck, J. A., et al., Cell 76:333-343 (1994); Weis, K., et al., Cell 76:345-358 (1994); Koken, M. H., et al., EMBO J. 13:1073-1083 (1994)), the currently prevailing hypothesis is that PML-RARxcex1 possesses altered transcriptional properties compared to PML or RARxcex1 and/or may act in a dominant-negative manner (Perez, A., et al., EMBO J. 12:3171-3182 (1993); de The, H., et al., Cell 66:675-684 (1991); Kastner, P., et al., EMBO J. 11:629-642 (1992)).
Acute promyelocytic leukemia (APL) is the prototype of a cancer treated by differentiation therapy using ATRA (Fenaux, P. et al., Semin Oncol. 24:92-102 (1997)). However, adjuvant chemotherapy which is required to improve tumor cell remission bears the inherent risk of therapy-induced ATRA resistance due to, for example, mutation oft he PML-RARxcex1 ligand binding domain. Such mutations are indeed frequently observed in relapsed patients (Imaizumi, M. et al., Blood 92:374-382 (1998)) and ATRA-resistant cell lines (Ruchaud, S. et al., Proc. Natl. Acad. Sci. U.S.A. 91:8428-8432 (1994); Shao, W. et al., Blood 89:4282-4289 (1997); Kizaki, M. et al., Blood 88:1824-1833 (1996); Robertson, K. A. et al., Blood 80:1885-1889 (1992); Kitamura, K. et al., Leukemia 11:1950-1956 (1997)).
Breast Cancer
Despite earlier detection and a lower size oft he primary tumors at the time of diagnosis (Nystrom, L. et al., Lancet 341:973-978 (1993); Fletcher, S. W. et al., J. Natl. Cancer Inst. 85:1644-1656 (1993)), associated metastases remain the major cause of breast cancer mortality (Frost, P. and Levin, R., Lancet 339:1458-1461 (1992)). The initial steps of transformation characterized by the malignant cell escape from normal cell cycle controls are driven by the expression of dominant oncogenes and/or the loss of tumor suppressor genes (Hunter, T. and Pines, J., Cell 79:573-582 (1994)).
Tumor progression can be considered as the ability of the malignant cells to leave the primary tumoral site and, after migration through lymphatic or blood vessels, to grow at a distance in host tissue and form a secondary tumor (Fidler, I. J., Cancer Res. 50:6130-6138 (1990); Liotta, L. et al., Cell 64:327-336 (1991)). Progression to metastasis is dependent not only upon transformation but also upon the outcome of a cascade of interactions between the malignant cells and the host cells/tissues. These interactions may reflect molecular modification of synthesis and/or of activity of different gene products both in malignant and host cells. Several genes involved in the control of tumoral progression have been identified and shown to be implicated in cell adhesion, extracellular matrix degradation, immune surveillance, growth factor synthesis and/or angiogenesis (reviewed in, Hart, I. R. and Saini, A., Lancet 339:1453-1461 (1992); Ponta, H. et al., B.B.A. 1198:1-10(1994); Bernstein, L. R. and Liotta, L. A., Curr. Opin. Oncol. 6:106-113 (1994); Brattain, M. G. et al., Curr. Opin. Oncol. 6:77-81 (1994); and Fidler, I. J. and Ellis, L. M., Cell 79:185-188 (1994)).
However, defining the mechanisms involved in the formation and growth of metastases is still a major challenge in breast cancer research (Rusciano, D. and Burger, M. M., BioEssays 14:185-194 (1992); Hoskins, K. and Weber, B. L., Current Opinion in Oncology 6:554-559 (1994)). The processes leading to the formation of metastases are complex (Fidler, I. J., Cancer Res. 50:6130-6138 (1990); Liotta, L. et al., Cell 64:327-336 (1991)), and identifying the related molecular events is thus critical for the selection of optimal treatments.
In one aspect, the invention is directed to a method of treating a hyperproliferative disease in a subject, the method comprising: (a) administering to the subject a pharmaceutically effective amount of a retinoid X receptor (RXR) agonist; and (b) administering to the subject a pharmaceutically effective amount of an agent which is capable of activating protein kinase A (PKA). The method can further comprise (c) administering to the subject a pharmaceutically effective amount of a retinoic acid receptor (RAR) agonist. The method can further comprise (d) administering to the subject a pharmaceutically effective amount of a cytokine, with or without a pharmaceutically effective amount of an RAR agonist.
Also provided is a kit useful for carrying out the method of treating a hyperproliferative disease.
In another aspect, the invention is directed to a method of inhibiting proliferation of breast cancer cells by administering an RXR agonist and an agent capable of activating protein kinase A. Breast cancer cell lines include, but are not limited to, T47D.
In another aspect, the invention is directed to a pharmaceutical composition comprising (axe2x80x2) a retinoid X receptor (RXR) agonist and (bxe2x80x2) an agent capable of activating protein kinase A (PKA). The composition can further comprise (cxe2x80x2) a retinoic acid receptor (RAR) agonist. The composition can further comprise (dxe2x80x2) a cytokine, with or without an RAR agonist (cxe2x80x2).
RXR agonists include, but are not limited to, the group consisting of 9-cis retinoic acid, bexarotene, 4-[1-[5,6-Dihydro-3,5,5-trimethyl-8-(1-methylethyl)-2-naphthalenyl]ethenyl]benzoic acid, and SR11237.
The agent capable of activating PKA can be a PKA agonist. PKA agonists include, but are not limited to, 8-bromo-cAMP, Sp-cAMPS, 8CPT-cAMP, dibutyryl-cAMP, Sp-5,6-DCl-cBiMPS, adenylate cyclase toxin, forskolin, L-858051, and Sp-8-pCPT-cGMPS. Alternatively, the agent can be a compound that increases cAMP level, either by stimulating cAMP synthesis or by inhibiting a phosphodiesterase. Compounds which increase cAMP synthesis include, but are not limited to, adenylate cyclase toxin, forskolin, and L-85 8051. Compounds that act as inhibitors of phosphodiesterases include, but are not limited to, RO 20-1724, Rolipram, Etazolate, and 3-isobutyl-1-methylxanthine (IBMX).
RAR agonists include RARxcex1, RARxcex2 and RARxcex3 agonists. Of course, an RAR agonist can be selective or specific for one or more of the RAR subtypes. RARxcex1 agonists include, but are not limited to, 9-cis retinoic acid, all-trans retinoic acid, 4-[[(2,3-Dihydro-1,1,3,3-tetramethyl-2-oxo-1H-inden-5-yl)carbonyl]amino] benzoic acid, AM-80, and AM-580.
Cytokines include, but are not limited to, a granulocyte colony-stimulating factor (G-CSF), granulocyte macrophage colony-stimulating factor (GM-CSF) and macrophage colony-stimulating factor (M-CSF).
In the method, steps (a)-(d) can be done concurrently, or in any order.
Hyperproliferative diseases can be, but are not limited to, cancer and psoriasis. Cancers include, but are not limited to, acute promyelocytic leukemia and breast cancer. The subject can be resistant to treatment with an RARxcex1 agonist alone.